More than one-third of women with epilepsy have a catamenial seizure pattern, in which seizures fluctuate with the menstrual cycle. Catamenial seizures are due, in part, to effects of estradiol and progesterone on the neural circuitry that supports seizures, such as in the hippocampus. Our research aims to understand how estradiol's effects on synapses in the hippocampus contribute to altered seizure patterns, with the goal of identifying targets for novel anti-seizure therapies. Preliminary studies point to regulation of presynaptic release at inhibitory synapses in the hippocampus as an important mode of estradiol action that has a profound impact on hippocampus-dependent seizures. We find that twenty-four hours of estradiol treatment has two simultaneous and opposing effects on seizures: it increases seizure susceptibility, and at the same time, decreases seizure severity. The estradiol-induced increase in seizure susceptibility appears to be related to suppression of GABA release from inhibitory synapses, whereas the decrease in seizure severity is due to enhanced release of the anti-convulsant neuropeptide, neuropeptide Y. Furthermore, we find that clusters of small vesicles and some large dense core vesicles in inhibitory presynaptic boutons are immunoreactive for estrogen receptors. This suggests that estradiol exerts its effects on release of GABA and neuropeptide Y by acting at the level of presynaptic boutons, possibly directly on vesicles themselves. In support of direct action on inhibitory synapses, we also have discovered that estradiol can acutely, i.e., within minutes, suppress inhibitory synaptic transmission in the hippocampus. Together, these studies suggest novel presynaptic mechanisms by which estradiol influences hippocampal physiology and hippocampus-dependent seizures. We will use a combination of whole-cell voltage clamp recording in brain slices from rats and mice, light and electron microscopic studies of synapses and vesicles, cell fractionation and immunoisolation of vesicles, and behavioral analyses of seizures to investigate how estradiol regulation of inhibitory synapses in the hippocampus influences seizure patterns. Our studies will be divided into 3 specific aims to investigate: 1) estradiol suppression of GABAergic synaptic inhibition;2) estradiol facilitation of neuropeptide Y release;and 3) how estradiol's effects on GABA and neuropeptide Y release influence seizure susceptibility and/or severity, both acutely and on a longer time scale. PUBLIC HEALTH RELEVANCE The proposed research will investigate novel mechanisms by which the hormone, estrogen, regulates the release of neurotransmitters and neuropeptides at synapses in the hippocampus, a key brain region in epilepsy. Understanding the mechanisms by which estrogen influences hippocampal synaptic function is important to identify cellular and molecular targets for novel anti-seizure therapies.